Sunlight inactivation of fecal bacteriophages and bacteria in sewage-polluted seawater.

Abstract

Sunlight inactivation rates of somatic coliphages, F-specific RNA bacteriophages (F-RNA phages), and fecal coliforms were compared in seven summer and three winter survival experiments. Experiments were conducted outdoors, using 300-liter 2% (vol/vol) sewage-seawater mixtures held in open-top chambers. Dark inactivation rates (k(D)s), measured from exponential survival curves in enclosed (control) chambers, were higher in summer (temperature range: 14 to 20 degrees C) than in winter (temperature range: 8 to 10 degrees C). Winter k(D)s were highest for fecal coliforms and lowest for F-RNA phages but were the same or similar for all three indicators in summer. Sunlight inactivation rates (k(S)), as a function of cumulative global solar radiation (insolation), were all higher than the k(D)s with a consistent k(S) ranking (from greatest to least) as follows: fecal coliforms, F-RNA phages, and somatic coliphages. Phage inactivation was exponential, but bacterial curves typically exhibited a shoulder. Phages from raw sewage exhibited k(S)s similar to those from waste stabilization pond effluent, but raw sewage fecal coliforms were inactivated faster than pond effluent fecal coliforms. In an experiment which included F-DNA phages and Bacteroides fragilis phages, the k(S) ranking (from greatest to least) was as follows: fecal coliforms, F-RNA phages, B. fragilis phages, F-DNA phages, and somatic coliphages. In a 2-day experiment which included enterococci, the initial concentration ranking (from greatest to least: fecal coliforms, enterococci, F-RNA phages, and somatic coliphages) was reversed during sunlight exposure, with only the phages remaining detectable by the end of day 2. Inactivation rates under different optical filters decreased with the increase in spectral cutoff wavelength (50% light transmission) and indicated that F-RNA phages and fecal coliforms are more susceptible than somatic coliphages to longer solar wavelengths, which predominate in seawater. The consistently superior survival of somatic coliphages in our experiments suggests that they warrant further consideration as fecal, and possibly viral, indicators in marine waters.

Spectral transmission curves for the optical filters used as chamber covers, including the spectral cutoff wavelength (λ50) for each filter (obtained by scanning the filter materials from 300 to 650 nm, against an air reference, with a model PU8800 spectrophotometer). Also shown is an indicative solar spectrum.

Variability associated with sample collection, transport, and assay. Shown are box plots of the counts obtained from 10 samples collected over a period of 4 min from each tank. The crosspieces of each box plot represent (from top to bottom), maximum, upper-quartile, median, lower-quartile, and minimum values. An outlier (open circle) is defined as a point whose value is either above the upper quartile by 1.5 times the interquartile distance or below the lower quartile by 1.5 times the interquartile distance. Laboratory-cultured B. fragilis phages were added to the sewage inoculum.

Inactivation in seawater, as a function of insolation, of somatic coliphages (■), F-RNA phages (◊), and fecal coliform bacteria (●) from untreated sewage (data from all survival experiments). The fecal coliform data are the linear portions of the inactivation curves (i.e., shoulder points, where present, have been removed). The R2 values are as follows: for somatic coliphages, 0.82; for F-RNA phages, 0.86; and for fecal coliforms, 0.89.

Inactivation in seawater, as a function of insolation, of somatic coliphages (■), F-RNA phages (◊), and fecal coliform bacteria (●) from raw sewage (A) and waste stabilization pond (WSP) effluent (B). For clarity, the phage data are presented as regression lines (the raw sewage R2 values are as follows: for somatic coliphages, 0.89; and for F-RNA phages, 0.90; the WSP effluent R2 values are as follows: for somatic coliphages, 0.96; and for F-RNA phages, 0.99).

Inactivation in seawater of somatic coliphages (■), F-RNA phages (◊), enterococci (○), and fecal coliform bacteria (●) from untreated sewage, as a function of insolation and time. The insolation scale is linear during daylight hours; the time scale is linear during the overnight period. Samplings in which no CFU were detected in 100 ml are presented as <1 on the (log10) y axis.